Methotrexate (MTX) is a potent antimetabolite used increasingly for a variety of medical indications, besides its established use for neoplastic disease. Because toxicity ir primarily determined by duration of exposure of vulnerable tissues to the drug, and since the drug is almost entirely excteted unchanged in the urine (by both filtration and active tubular secretion), any condition which delays renal excretion is likely to increase toxicity. The weak organic and acid drugs probenecid and salicylate have been shown to inhibit the tubular secretion of MTX to a clinically important degree. However, it is not known which other weak organic acid drugs are capable of similarly important inhibition. Case reports and personal observations suggest that several other weak organic acid drugs have caused serious or fatal drug interactions when given with low doses of MTX. The long-term objective of this project is to determine which weak organic acid drugs are capable of inhibiting the active tubular secretion of MTX to a clinically important extent, thereby predisposing to MTX toxicity. To achieve this goal, a research plan with five stages has been developed. First, the rabbit kidney slice model of tubular secretion will be used to measure the active accumulation of MTX, p-aminohippurate (PAH), and penicillin G (PEN). The ability of other weak organic acid drugs to inhibit this active accumulation will be investigated as well. Second, this model will be extended to investigate the nature of such inhibition when it occurs. When the inhibition is found to be competitive in nature, the drug inhibitor-carrier protein dissociation constant (Ki) will be calculated. Such calculations will generate evidence concaerning a common secretory pathway for MTX, PAH, and PEN. Third, the therapeutic plasma concentration and Ki value for each inhibitory drug will be compared, and these calculations used to predict which organic acid drugs would be most likely to inhibit the tubular secretion (and thus renal clearance) of MTX and PEN in vivo. Fourth, these predictions will be compared to the known clinical effects of four drugs upon MTX renal clearance, and the known clinical effects of nine drugs upon PEN renal clearance. Fifth, the predictions of the in vitro model will be further tested by studying the effects of organic acid drugs upon MTX renal clearance in patients already receiving MTX and potential drug inhibitors. The information gained should allow MTX to be used in a more rational and less toxic manner.